Evidence for a nitrate-independent function of the nitrate sensor NRT1.1 in Arabidopsis thaliana

Abstract

NRT1.1 is a putative nitrate sensor and is involved in many nitrate-dependent responses. On the other hand, a nitrate-independent function of NRT1.1 has been implied, but the clear-cut evidence is unknown. We found that NRT1.1 mutants showed enhanced tolerance to concentrated ammonium as sole N source in Arabidopsis thaliana. This unique phenotype was not observed in mutants of NLP7, which has been suggested to play a role in the nitrate-dependent signaling pathway. Our real-time PCR analysis, and evidence from a literature survey revealed that several genes relevant to the aliphatic glucosinolate-biosynthetic pathway were regulated via a nitrate-independent signal from NRT1.1. When taken together, the present study strongly suggests the existence of a nitrate-independent function of NRT1.1.

Keywords

Ammonium tolerance Ammonium toxicity Nitrate sensor NRT1.1

Electronic supplementary material

The online version of this article (doi:10.1007/s10265-010-0385-7) contains supplementary material, which is available to authorized users.

Notes

Acknowledgments

We thank Dr. M. Taira, Dr. H. Hirano, Dr. Y. Ohmori and Dr. Y. Yasuoka for the use of the real-time PCR system. This study was financially supported by the Sasakawa Scientific Research Grant from The Japan Science Society, and by the Ministry of Education, Science, Sports, and Culture (Nos. 1685101, 17770015).

Supplementary material

Supplementary Table S1 List of genes whose expression levels were changed more than three fold by the mutation of NRT1.1 in the absence of nitrate (in the presence of chloride). Red bold letters denote aliphatic glucosinolate-biosynthetic genes. This list was based on the supplemental array data of Wang et al. (2009) with some modifications. See the original manuscript for other details on experimental procedures (PDF 71 kb)

Supplementary Figure S1 (a) Genomic PCR using a pair of NRT1.1-specific primers (NRT1.1F, NRT1.1R) and a T-DNA-specific primer (LB). A PCR product (ca. 2600 bp) with NRT1.1F and NRT1.1R was amplified from Col-0 (c), but not from SALK-097431 (n). A PCR product (ca. 900 bp) with NRT1.1F and LB was amplified from SALK-097431, but not from Col-0. The putative position of the T-DNA insertion in the NRT1.1 gene of SALK_097431 is shown. Boxes correspond to exons. (b) Real-time PCR and RT-PCR using a pair of NRT1.1-specific primers (CHL1F, CHL1R; Hu et al. 2009). Relative transcript level of NRT1.1 mRNA in SALK-097431 was less than 1% of Col-0 (PDF 35 kb)

Supplementary Figure S2 Effects of different nitrogen and pH conditions on the growth of A. thaliana. In Figure S2b, seeds of Col-0 and nrt1.1, or Col-0 and chl1-5 were placed on the same 10A-medium. One dish contained seven seeds per line. The dishes were set in a vertical position, and the plants were grown for eleven days. See the legend of Figure 1 for more detailed information (PDF 225 kb)

Supplementary Figure S3 Transcript levels of glucosinolate-biosynthetic genes in the roots of Col-0- and nrg1 (a NRT1.1 mutant). Graphs are presented using the mean signal intensity in Table S1. See the original manuscript of Wang et al. (2009) for other details on experimental procedures (PDF 56.6 kb)